Neutrophil elastase (NE) decreases G-CSFR expression on human PMN. Peripheral blood neutrophils (PMN) from healthy donors (97% purity) were incubated (1 × 107 cells/ml) with 150 μg/ml NE at 37°C for the indicated times. G-CSFR surface expression was analyzed by flow cytometry as described using a biotinylated anti-G-CSFR antibody recognizing FNIII domains in the extracellular region of the G-CSFR (BD PharMingen) and a BD FACSCalibur Cytometer. A representative experiment from five independent experiments using neutrophils from five different donors is shown. (A) Effect of NE treatment for the indicated times on G-CSFR expression on PMN. Cells incubated with an isotype-matched control antibody (Iso) are shown as a negative control. (B) Effect of inclusion of 10% FBS (+) during incubation of PMN with NE. (C) PMN were treated with 150 μg/ml of NE, azurocdin (AZ), or cathepsin G (CG) for 2 h, then the G-CSFR was analyzed by flow cytometry. Shown is percent of fluorescence compared to maximum fluorescence at time 0.